Biological systems and the processes which operate therein or are caused to so operate have their basic foundation on the interaction of molecules. Molecular forces in biological systems are different from other molecular systems where chemical reactions and changes of physical phases in a complete system are concerned. Statements about molecular interactive activity in biological systems bring forward the advantageous concept that such systems could be analyzed in order to be able in the future to formulate statements in greater detail.
For the measurement of molecular interchange in biological systems, among other methods, raster type scanning, based on probe centered, microscopic methods are in frequent use, in order to make determinations regarding the topographic characteristics of surfaces by using high lateral and vertical resolution. In regard to “lateral resolution”, is to be understood that resolution in a single plane of a surface under investigation in a biological system is meant, while correspondingly, the resolution perpendicularly aligned thereto is designated as “vertical resolution”.
Examples for raster-scanning probe microscopic applications encompass such procedures as may be found in, for instance, SFM (scanning force microscopy) or again in AFM (atomic-force microscopy).
With such scanning probe microscopy applications, it is possible, that besides the topology of a surface of a biological sample, also the elasticity thereof or the thereto applicable adhesion forces can be determined. The scanning probe microscopy, here normally referred to as a “force-spectroscopy”, determines molecular forces of the sample by means of an analytic probe which invades or reacts with the said sample, in order to quantitatively detect and characterize the interactive exchange between individual molecules. Customarily, a probe includes a measuring tip, which is carried on a freely extending arm or is supported in a cantilever fashion. When being used in an analysis, the probe assembly is measurably caused to crisscross over the surface of a sample, during which action, the lateral and vertical positions and/or deflections of a probe can be recorded. Movements of a probe relative to the sample are possible due to the elasticity of a probe itself and especially the characteristics of the said cantilever holder thereof. Based on lateral and vertical positioning and/or deviations of a probe, molecular forces inherent in the sample and the therefrom induced topographical characteristics can be determined.
Customarily, movements of a probe are detected by optical measuring instruments, wherein the resolution stands at about 0.1 nm and permit a determination of forces in the range of a few pN (pico-Newton).
In order to determine the surface topography of a biological sample, the surface of the sample and the probe of the said force-microscopic procedure can be so mutually counter positioned, that a force, acting between them can be brought to a predetermined value (nominally about 50 to 100 pN). Accordingly, a probe and the sample are so placed, relative to one another, that a crisscross (raster) scan of the surface of the sample is enabled. In this way, the sample and/or the probe can move also vertically, so that the forces acting between them are purposely retained within the predetermined values. Movements of the sample and probe relative to one another can be activated by means of a piezoelectric ceramic device.
One advantage of the scanning force-microscopy can be found therein, in that biological samples in buffered solutions held at a relevant physiological temperature (namely 4° C. and 60° C.) can be investigated.
FIG. 1 illustrates, in a simplified way, the principal of a scanning force microscope. For the analysis of the surface of a biological sample BP a probe S is employed. A probe S can be considered to be a probe-tip Sp, which is suspended from a spring C. In a case of movement of a probe S and the biological sample BP relative to one another, (for example along the path W), dependent upon the respective surface topography, a probe S and the sample SP are so mutually displaced in relation to one another in a vertically designated direction, that the displacement of the spring C remains constant.
At the present time, scanning probe microscopy, however, presents a task which is time consuming and intensive of personal attention. Actually, a great number of measurement increments, are necessary for a well founded analysis. For this reason, present applications of scanning probe microscopy are entirely unsatisfactory, due to the small number of measurements made, for instance, per day. Also the limited possibilities of the analyses, of the evaluation and the subsequent use of the measured data, present considerable disadvantages.